Retroviral-mediate gene therapy efforts inhuman have been plagued by the loss of therapeutic gene expression, due to factors such as inactivation of viral vector promoters. Native promoters and newly characterized insulator sequences offer promising solutions to this challenge. We have achieved short-term gene correction in vivo using Lamellar Ichthyosis (LI) as a prototype human genetic skin disease. Our next step is to sustain this correction based on the hypothesis that new genetic regulatory elements will generate sustained corrective gene delivery. To do this we will utilize endogenous human promoters active in the epidermis to generate and test novel insulated high titer amphotrophic retroviral gene delivery vehicles. Because an entirely murine model may be of limiter future clinical relevance due to the major differences between human and mouse and mouse epidermal structure and biology, we utilize normal and diseased human keratinocytes to develop human epidermal gene delivery. TO do this, we will utilize the severe combined immunodeficiency (SCID) mouse xenograft model where grafts may be used to regenerate human skin on immunodeficient mice. The long term objective of this proposal is to develop effective approaches to sustainable therapeutic cutaneous gene delivery, using LI as a human disease model, as a foundation for molecular therapy of genetic skin disease. Based on the hypothesis that LI keratinocytes-corrected via optimized sustainable vectors-will give rise to a human epidermis with a sustained normal phenotype, we will develop a model of human cutaneous gene therapy in vivo.